Visceral leishmaniasis is a potentially fatal disease which affects individuals in many tropical and subtropical countries. There is an ongoing outbreak of the disease, cause by the protozoan Leishmania donovani chagasi, in the State of Rio Grande do Norte in Northeast Brazil. Currently there is no proven form of immunoprophylaxis against the disease. However, recovery from visceral leishmaniasis usually results in long-term immunity against reinfection, suggesting that antigens inducing an immunoprotective response exist. The purpose of this proposal is to identify and characterize parasite antigens that stimulate immune responses in human hosts that are immune to the disease. There is considerable evidence indicating that cellular immune responses are critical for both recovery from, and subsequent immunity to reinfection with L. d. chagasi. According to clinical criteria, a subset of humans living in an endemic region that are immune can be identified. Their peripheral blood lymphocytes respond in an antigen-specific manner to parasite antigens, and at least some of these antigens are likely to be responsible for the immunoprotective response. identification of these antigens would therefore be of tremendous value in achieving our goal. Within a mammalian host Leishmania survive in macrophages exclusively in the form of obligate intracellular amastigotes. Therefore parasite- specific T cell responses are likely to be directed against amastigote antigens. Amastigote proteins, however, are difficult t study because of their small size and obligate intracellular location. Therefore we have constructed a cDNA expression library from amastigote RNA, to use as a source of amastigote proteins. These recombinant proteins, as well as several native parasite antigens, will be used to test responses of peripheral blood lymphocytes from humans that are found to be immune to the disease according to clinical criteria. This collaborative proposal takes advantage of the ability of the ability of the Brazilian investigator to study hosts who are immune to visceral leishmaniasis, and the availability of molecular and immunologic techniques to the U.S. investigator. The goal of the project is to identify antigens that stimulate parasite-specific responses in immune human lymphocytes. The strategy takes advantage of the ease and efficiency of recombinant DNA techniques. These studies are aimed at identifying antigens that might eventually be useful in developing a protective vaccine.